Hyperthermia is one of the most effective radiosensitizers known. Therefore, the goal of Project 1 is to delineate the mechanisms of radiosensitization by short-duration moderate hyperthermia and the mechanism(s) by which radiosensitization is increased via increasing thermal dose, so that the cellular targets for enhancers of heat-induced radiosensitization will be defined. It is known that hyperthermia enhances protein binding to nuclear components, leading to altered protein-protein interactions throughout the nucleus. Accumulating evidence suggests that some of these changes cause radiosensitization. However, the exact changes in nuclear protein interactions that contribute to radiosensitization remain unknown. Thus, the objectives of the proposed work are to delineate the changes in nuclear protein associations induced by short duration, moderate hyperthermia that lead to increased radiosensitivity. The first step will be to determine if the heat-induced translocation of the DNA-repair protein, Mrel 1, from the nucleus to the cytoplasm and/or its dissociation from its functional partners causes radiosensitization by short-duration, moderate hyperthermia (Specific Aim 1). Second, we will determine which changes in nuclear protein association contribute to increasing radiosensitization by increasing thermal dose. The second part of the hypothesis is that the heat-induced increase in the binding of proteins to DNA-nuclear matrix attachment regions contribute to increased radiosensitivity at thermal doses above the minimum required for radiosensitization (e.g., acute hyperthermia). If altered protein conformations are responsible for changes in protein associations that lead to heat-induced radiosensitization, it is likely that recovery from these effects would require an association between the protein and a molecular chaperone(s) e.g., hsp70, which will be tested in Specific Aim 3. The hypothesis that an increasing spectrum of radiosensitizing effects contribute to enhanced radiosensitization will be tested in Specific Aim 4, by determining if moderate hyperthermia, plus an enhancer of HIR produces at least some of the radiosensitizing effects of acute hyperthermia.